Biochemical studies on antiparasitic drugs.
Item
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Title
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Biochemical studies on antiparasitic drugs.
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Identifier
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AAI9707104
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identifier
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9707104
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Creator
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Hong, Yu-Long.
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Contributor
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Adviser: Steven R. Meshnick
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Date
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1996
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Language
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English
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Publisher
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City University of New York.
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Subject
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Chemistry, Biochemistry | Health Sciences, Pharmacology
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Abstract
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Understanding the mechanisms of antiparasitic drugs is necessary for the development of new drugs. The modes of action of sulfa drugs and artemisinin are studied in this thesis.;P. carinii synthesizes folates de novo from exogenous p-aminobenzoic acid (pABA). The predominant forms of folates in P. carinii are pteroylpentaglutamates. Culture-derived P. carinii produces pteroylpentaglutamates at a four-fold higher specific activity than lung-derived organisms do, possibly because they contain less contaminating lung debris.;Forty-four sulfa drugs were screened against crude preparations of recombinant P. carinii dihydropteroate synthetase (DHPS). The apparent Michaelis-Menten constants (K{dollar}\sb{lcub}\rm m{rcub}{dollar}) for p-aminobenzoic acid and 7,8-dihydro-6-hydromethylpterin pyrophosphate are 0.34 {dollar}\pm{dollar} 0.02 and 25 {dollar}\pm{dollar} 0.71 {dollar}\mu{dollar}M, respectively.;Several sulfa drugs inhibit DHPS approximately as well as sulfamethoxazole, as determined by the concentrations which cause 50% inhibition and/or by K{dollar}\sb{lcub}\rm i{rcub}{dollar} values. For all sulfones and sulfonamides tested, unsubstituted p-amino groups are necessary for activity, and sulfonamides containing N1-heterocyclic substituent are found to be the most effective inhibitors.;Folate biosynthesis in isolated intact P. carinii is approximately equally sensitive to inhibition by sulfamethoxazole, sulfathiazole, sulfachlorpyridazine, sulfamethoxypyridazine, and sulfisoxazole. Two of these drugs, sulfamethoxypyridazine and sulfisoxazole, are known to be less toxic than sulfamethoxazole and should be further evaluated for the treatment of Pneumocystis carinii pneumonia (PCP).;The recombinant P. carinii Fas was expressed as a glutathione S-transferase fusion protein in baculovirus-infected insect cells, and then purified by affinity chromatography using glutathione beads. The overall yield of DHPS activity was 4.6% with a purification of 39-fold.;In vitro, hemin and artemisinin were found to undergo a chemical reaction forming two major products which were isolated by high-performance liquid chromatography (HPLC). The m/z's of the two products were 856 and 871.;({dollar}\sp{14}{dollar}C) -Artemisinin was taken up by Plasmodium falciparum in culture and concentrated in hemozoin. Majority of the hemozoin-associated radioactivity comigrated with the synthetic adducts.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
